Metabolically Active Macrophages: A novel mechanistic link between obesity and TNBC Triple negative breast cancer (TNBC) patients have an extremely poor prognosis due to their high metastatic potential and lack of targeted drug therapies. Emerging epidemiological data suggest that obesity is strongly linked to the incidence and severity of TNBC. Thus, understanding the biological processes that link obesity and TNBC has important clinical applications for prognosis and treatment. Previous study showed that obesity promote tumor burden in C3Tag mouse model. However, mechanisms by which obesity potentiates TNBC progression are incompletely understood. One clue to its action is that obesity causes chronic inflammation, and adipose macrophages (ATMs) accumulation, which are a key source of this inflammation. While it is well established that tumor-associated macrophages (TAMs) are key effector cells that promote breast cancer, role of ATMs on tumorigenesis has largely been ignored. ATMs are an attractive mechanistic link between obesity and TNBC as ATMs are the predominant type of macrophage in the breast during early tumorigenesis. This raises the possibility that ATMs play an important role in tumor progression. Here we show that metabolic dysfunction promotes a mechanistically distinct pro-inflammatory phenotype (metabolic activation; MMe) in breast adipose tissue macrophages isolated from obese women and express cell surface markers of MMe (CD36, ABCA1), but not M1 (CD38, CD319, CD274), macrophages. Furthermore, treating nave macrophages with media conditioned by human breast adipose tissue from an obese subject (BMI = 37), but not a lean subject (BMI = 19), induced genes diagnostic of the MMe phenotype. We further demonstrated that pre-treating human TNBC cells (MDA-MB-231) with conditioned media derived from MMe macrophages promotes colony formation and invasion in vitro, and intravasation of cancer cells into the blood in vivo. Moreover, pre-treated cancer cells showed a two fold increase in the expression of stem-like cell markers (CD90, OCT4, SOX2 and NANOG). Remarkably, we saw the same effect on ?stemness? of tumor cells in obese mice compared to lean mice. Based on these findings we hypothesize that obesity-induced changes to mammary adipose tissue reprogram macrophages to a MMe phenotype that potentiates TNBC initiation and metastasis. In the proposed work, we would determine the mechanism by which MMe macrophages promote TNBC progression (aim1) and their contribution in promoting obesity-associated TNBC (aim 2).
In aim 1, we will test the hypothesis that inflammatory cytokines secreted by MMe macrophages are responsible for their pro- tumorigenic effect. Two test this we will deplete inflammatory cytokines in MMe macrophage conditioned media using two approaches: i). Genetic knockout (TLR2-/- and NOX2-/-) macrophages that inhibit inflammatory cytokine expression in MMe macrophages. ii). Neutralizing antibodies against inflammatory cytokines. We would then conduct various in vitro and in vivo functional assays to test the effect of MMe macrophage conditioned media on tumor cell phenotype.
In Aim 2, we hypothesize that mammary ATMs action during early tumorigenesis is required for obesity induced TNBC progression. To test this, we will use clodronate liposomes to ablate ATMs from mammary adipose tissue of obese mice before injecting cancer cells in a novel syngeneic mouse model (REAR mice). We will monitor the tumor progression and metastasis using live mice xenogen imaging. We would then isolate these tumor cells to test expression of stem-like cell markers and initiation using limited dilution assays. In our knowledge this is the first study focused on understanding the role of mammary ATMs in obesity- associated TNBC progression. A comprehensive understanding of the signaling mechanism(s) involved in metabolic activation of mammary ATMs would enable development of directed therapies towards this specific pro-tumorigenic macrophage phenotype, thereby leaving the immune system of cancer patients intact.
Triple negative breast cancer (TNBC) accounts for about 20% of all breast cancers. TNBC patients have an extremely poor prognosis due to their high metastatic potential and lack of targeted drug therapies. Emerging epidemiological data suggest that obesity is strongly linked to the incidence and severity of TNBC; obese women have a 35% higher risk of developing TNBC and 46% higher risk of developing distant metastases. However, the role of obesity in TNBC progression is largely unexplored. Thus, understanding the biological processes that link obesity and TNBC has important clinical applications for prognosis and treatment.
